Objective To investigate the effects of Buyang Huanwu decoction and its decomposed recipes on neurological function and angiogenesis after focal cerebral ischemia in rats and its mechanism. Methods Fifty-two clean grade SD male rats were randomly divided into sham-operation,model,whole prescription,invigorating qi,and promoting blood circulation groups (n = 8 in each group)according to the random number table. In addition to the sham-operation group,the middle cerebral artery occlusion models of the rats in other groups were induced by the suture method. The patients with the first Longa nerve function scores 1 to 3 were used as successful modeling. The whole Buyang Huanwu decoction included dried root of Astragalus membranaceus 120. 0 g,dried root of angelica sinensis 6. 0 g,dried root of Paeonia lactiflora 4. 5 g, dried rhizome Ligusticum chuanxiong 3. 0 g,dried body of Pheretima aspergillum 3. 0 g,dried flowers of Carthamus tinctorius 3. 0 g,and seed of Prunus persica 3. 0 g;the invigorating qi prescription included dried root of Astragalus membranaceus 120. 0 g;the promoting blood circulation prescription included dried root of angelica sinensis 6. 0 g,dried root of Paeonia lactiflora 4. 5 g,dried rhizome Ligusticum chuanxiong 3. 0 g, dried body of Pheretima aspergillum 3. 0 g,dried flowers of Carthamus tinctorius 3. 0 g,and seed of Prunus persica 3.0 g. On the first day after procedure,the rats began to be administered intragastrically. The intragastric doses of the whole prescription group,invigorating qi group,and promoting blood circulation group were 13. 1,10. 8,and 2. 2 g/ kg,respectively. The sham-operation group and the model group were given equal volume of isotonic saline,once a day for 14 days. 5-bromodeoxyuridine (BrdU,50 mg/ kg)were injected intraperitoneally,once a day for 14 days. The modified neurological severity score (mNSS)and the corner test were used to evaluate sensorimotor function at day 1,7 and 14 after procedure. BrdU and rat von Willebrand factor (vWF)double immunofluorescent staining were used to detect the angiogenesis in ischemic peripheral region;Western Blot was used to detect the protein expression of vascular endothelial growth factor (VEGF). Results (1)Compared with the model group,the mNSS score in rats in the whole prescription group was lower at day 7 and 14 after procedure (6. 8 ±1. 0 vs. 8. 5 ±1. 1,6. 1 ± 0. 8 vs. 8. 0 ± 1. 4;all P < 0. 01). The number of turning right in the whole prescription group was reduced (7. 1 ±0. 6 vs. 8. 6 ±1. 2 and 6. 1 ± 0. 8 vs. 7. 9 ±1. 1;all P < 0. 01). The number of turning right in the invigorating qi group was reduced (7. 5 ± 0. 5 vs. 8. 6 ± 1. 2 and 6. 2 ± 1. 0 vs. 7. 9 ± 1. 1;all P < 0. 01). At day 14 after procedure,the number of BrdU / vWF co-labeled immunopositive cells in ischemic peripheral zone of the whole prescription group was increased significantly. There was significant difference between the groups (30 ± 8 / mm2 vs. 24 ± 7 / mm2;P < 0. 01). The VEGF protein expression was increased (0. 33 ±0. 01 vs. 0. 30 ±0. 01;P <0. 01). (2)Compared with the invigorating qi group,the rat mNSS scores of the whole prescription group were lower at day 7 and 14 after procedure (the invigorating qi group 8.2 ±1.3 and 7.5 ±0.9 respectively;all P <0. 05). The number of BrdU/ vWF immunopositive cells in the whole prescription group was increased at day 14 after procedure (26 ±5/ mm2 in the invigorating qi group;P < 0. 05). The VEGF protein expression was increased (0.31 ±0.01 in the invigorating qi group;P <0.01). (3)Compared with the promoting blood circulation group, the mNSS scores of the whole prescription group were lower at day 7 and 14 after procedure (the promoting blood circulation group 8.5 ±0.9 and 7.6 ±0.7 respectively;all P <0. 05). The number of turning right was reduced (8.5 ±0. 8 and 7. 6 ± 0. 9 respectively in the promoting blood circulation group;all P < 0. 05). The number of BrdU/ vWF immunopositive cells in ischemic peripheral zone of the whole prescription group at day 14 after procedure was increased (26 ± 6 / mm2 ,P < 0. 05). The relative expression level of VEGF was increased (0. 31 ±0. 01 in the promoting blood circulation group,P <0. 05). Conclusion Buyang Huanwu decoction can promote angiogenesis and recovery of neurological function after cerebral ischemia. Its mechanism may be associated with the up-regulation of the VEGF protein. The traditional Chinese medicines for invigorating qi and invigorating the circulation of blood in the prescription have synergistic effect.

Objective To analyze the expression of hsa-miR-10b in three cervical cancer cell lines , and to find the target genes of hsa-miR-10b.Methods PCR was applied to measure expression levels of hsa-miR-10b in C-33A, HeLa and CaSki .The relative studies on hsa-miR-10b were retrieved from PubMed .The sequence and genome char-acteristics of hsa-miR-10b were analyzed on line by miRbase and NCBI .The target genes of hsa-miR-10b were searched by TargetScan , PicTar and miRanda , and then demonstrated by Gene Ontology and Pathway Enrichment analysis.Results Compared with C-33A, the expression of hsa-miR-10b significantly reduced in HeLa and Caski (P<0.01).Current studies showed that hsa-miR-10b was related with multiple tumorigenesis .hsa-miR-10b was lo-cated in human chromosome 2q31.1 and highly conserved among different species .The target genes were enriched in the biological processes of transcription , gene expression regulation , cell proliferation and etc .(P<0.05).Pathway analysis showed that these target genes were responsible for biochemical erents mediated by to the signaling pathways of cancer, cell cycle, ARF and etc.(P<0.05).Conclusions hsa-miR-10b may have extensive functions , and closely related with the occurrence and development in cervical cancer .Prediction of target genes provides a theoreti-cal basis for the further study .

Objective To investigate the effect of Buyanghuanwu decoction(BYHWD) on the expression of growth-associated protein 43(GAP-43) and synaptophysin(SYP) after focal cerebral ischemia in mice.Methods Focal cerebral ischemia was induced by 30 min of middle cerebral artery occlusion followed by reperfusion.BYHWD (20 g/kg) was administered orally 24 h after ischemia and once a day.The neurological score and the corner test were used to evaluate sensorimotor function on 1,3,7,and 14 days after ischemia.The expression of GAP-43 and SYP in the ischemic boundary zone was determined by immunohistochemistry 14 days after ischemia.Results Compared with the model group,BYHWD significantly ameliorated neurological dysfunction at 3,7 and 14 days and reduced the number of right turn at 7 and 14 days after ischemia(P ＜ 0.05).The average optical density of GAP-43 and SYP was 0.217 ±0.012 and 0.278 ±0.019 at the boundary zone in model group 14 days after ischemia,respectively.BYHWD also significantly increased the immunoreactivity of GAP-43 (0.250 ± 0.013)and SYP(0.323 ± 0.017) (P＜ 0.01).Conclusion BYHWD promotes axonal regeneration and synaptic plasticity in the ischemic boundary zone,which may contribute to functional recovery after focal cerebral ischemia.

AIM: To study the effect of Liuwei Dihuang Wan (LWDHW) on the expression of immune-associated genes in the aged rats using cDNA microarray. METHODS: Forty 20-months-old rats were divided into two groups equally. One group was treated with LWDHW for 5 weeks,another was untreated as control. Some biochemical assays were used to confirm the physiological differences between two groups. The mRNA from the spleens of twenty 4-months-old young rats were extracted and divided equally into two parts. Each part was conjoined with mRNA from drug treated aged rats and that from untreated control aged rats,respectively,and two experimental combinations for reverse transcription and cDNA microarray were formed. By comparing two groups of data from two combinations,the effects of LWDHW on the expression of the associated genes were evaluated. RESULTS: 13 down-regulated genes and one up-regulated gene were identified in the untreated control old rats,whose expression did not change obviously in treated old rats compared with young rats. The expressions of another two sequence were up-regulated distinctly in treated old rats compared with young rats. CONCLUSIONS: 16 immune-associated genes expressions were affected markedly by LWDHW. It will be helpful for understanding the molecular mechanism of LWDHW in ageing.

Objective To observe the effects of different levels manganese (Mn) on spatial learning and memory in neonate rats.Methods Neonate rats were distributed to control (normal saline) and MnCl210,20,30mg/kg groups randomly.Each groups included 10 litters in a cage with a dam.Neonate rats were intraperitoneal injection exposed to MnCl2 over PND 1-21.All groups were evaluated behavioral performance using open field and Morris water maze.Blood and hippocampus Mn levels were determined using ICP-MS.Results 1) For each group,blood Mn were (35.58 ± 13.77) μg/L,(80.00 ± 12.98) μg/L,(238.51 ± 31.43) μg/L,(348.47 ±34.07) μg/L and hippocampus Mn were (576.82 ± 79.78) μg/g,(798.33 ± 40.60) μg/g,(1017.23 ± 117.23)μg/g,(1278.76 ± 281.48) μg/g respectively.Blood and hippocampus Mn concentrations in Mn-exposed groups were significant increased compared to control (P ＜ 0.01),and there was a positive correlation in blood Mn and hippocampus Mn(OR =0.91,95％ CI=0.81-0.96,P＜ 0.01).2) Therewere no significant differences on travelled distance in open field among all groups,which meant that Mn exposure had no effect on their locomotion.3) In the hidden platform trials of the Morris water maze test,only on 3rd day,Mn-expose groups spent more time to find the platform compared to the control(P ＜ 0.01).The average escape latency were(21.77 ± 7.10)s,(33.78 ± 9.95)s,(37.17 ± 13.68) s,(41.92 ± 16.74) s respectively.Though the latency were increased with the Mn exposure levels increasing among the Mn-expose groups,no statistically significant differences were observed.There were no statistically effects on latency to find the platform of all groups in other training days.The result in probe trails showed that there were no statistically effects on swimming velocity,the number of crossing over the former platform and the time spent in the targeted quadrant.Conclusion Mn exposure exerts effects on the learning,but no doseeffect relationship.There are no effects on memory of neonate rats of Mn exposure.

BACKGROUND: Previous studies have confirmed that embryonic stem cells call be induced to differentiate into insulin-producing cells, but the induction process takes a long time. Most of the processes take about one month.OBJECTIVE: Activin A, all-trans retinoic acid (ATRA), basic fibroblast growth factor (bFGF) and nicotinamide were applied in vitro in combination to observe whether mouse embryonic stem cells could be induct to differentiate into insulin-producing cell in a relatively short time.DESIGN: Cell observation experiment.SETTING: Shanghai Institute of Endocrine and Metabolic Diseases, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.MATERIALS: This study was performed at Shanghai Institute of Endocrine and Metabolic Diseases from October 2004 to February 2006. Two mice of clean grade and of 12.5-14.5 days of gestational age were provided by Shanghai SLAC Laboratory Animal Co., Ltd (Permission No. 2004A034). The protocol was performed in accordance with ethical guidelines for the use and care of animals. Mouse embryonic stem cell lines were supplied by Dr Changxian Zhang (CNRS UMR5641, France). Activin A was the product of the R & D Corporation. ATRA and nicotinamide were supplied by the Sigma Corporation, USA. BFGF was supplied by Gibco Corporaion. METHODS: Head and viscera were removed from embryos of the pregnant mouse. The remaining tissues were cut into pieces and digested with trypsin. Cell suspension was centrifuged and inoculated at 3×108L-1. The cells could be used as mouse feeder layer after 2-3 times of passage. The mouse embryonic stem cells (ESCs) were inoculated onto the feeder layer in knockout Dulbecco's modified Eagle medium (DMEM) supplemented with leukemia inhibitory factors (LIF). ESCs were passaged at 1:3-1:6 after 2-3 days of culture. Culture medium with serum was added into the culture dishes to terminate the digestion. Cell fluid was centrifuged and supernatant was discarded. The sediments were prepared into suspension and inoculated at 2.5×104 with LIF-free culture medium. After 24-48 hours, embryonic bodies (EBs) were collected and replated in 1% Matrigel-coated dishes. When began to adhere to the dishes, EBs were cultured in 10% FBS/DMEM supplemented with 100μg/L activin A for 24 hours. Then EBs were switched to 10% FBS/DMEM for 6-8 hours as an interval. After this interval. EBs differentiated were cultured in 10% FBS/DMEM with 10<-6mol/L RA for another 24 hours followed by culture in 10% FBS/DMEM supplemented with 10μg/L bFGFs for 3-5 days. Finally, EBs differentiated were cultured in DMEM/F12 supplemented with N2 supplement, B27 supplement, 1μg/L laminin, 10μg/L bFGFs, and 10mmol/L nicotinamide for 3-5 days. Dithizone (DTZ) staining, inununofluorescent staining and reverse transcription-polymerase chain reaction (RT-PCR) were applied to detect insulin expression in the differentiated cells.MAIN OUTCOME MEASURES: Induction of ESCs, DTZ staining and immunofluorescent staining as wel as RT-PCR detection.RESULTS: Mouse ESCs growing on a feeder layer formed many colonies with clear boundary and dense structure. However, there was no obvious outer limit between these ESCs. EBs began to adhere to the dishes, which were coated with matrigel, on the 2nd day. After activin A and ATRA interval induction, EBs spread, and most of the living cells were epithelial cell-like when cultured in 10% FBS/DMEM supplemented with 10μg/L bFGFs. After culturing in DMEM/F12 supplemented with N2, B27, nicotinamide, bFGFs and laminin, the cells formed small clusters. The insulin-producing cells were stained dark red with DTZ, and the cells stained with primary antibody to insulin were insulin-positive. After 2 weeks of induction of activin A, ATRA, bFGFs and nicotinamide, the insulin-producing cells expressed insulin 2, Pdxl, Nkx6.1, Nkx2.2, PP, IAPP, Glut2, Somastatin, Hnf3β and Neuro D mRNA but did not express insulin 1 mRNA.CONCLUSION: Mouse ESCs call be induced to differentiate into insulin-producing cells by activin A, ATRA, bFGFs and nicotinamide in vitro. Induction time call be shortened to 2 weeks.

Objective To investigate the effect of bibliotherapy in soothing the postoperative pain in children and relieving the perioperative anxiety of children and their care givers. Methods Hospitalized children and their care givers from August 2007 to March 2009 were studied. 153 cases from August 2007 to May 2008 were assigned to the control group and the other 153 cases to the intervention group. Routine surgical nursing were applicated in the control group by introduction of perioperative nursing procedures.Bibliotherapy were applicated in the intervention group on the basis of the control group-using "bibliotherapy materials for hospitalized children" which was designed by ourselves and correspond with the theory of bibliotherapy to interfere in the 153 cases in the intervention group. The variance of preoperative anxiety of care givers and perceptions of postoperative pain of children between the two groups were compared with scales of mYPAS, STAI and FLACC and Wong- Baker Facial Scale. Results The scores of mYPAS in children of the intervention group and the control group was (35.875+4.441)and(46.796+8.606 )respectively and the variance was significant. The scores of STAI in care givers of the two groups was(38.125+4.371 )and (49.901 +7.420) respectively and revealed significant variance. The scores of Wong-Baker and FLACC in children of both groups 1 hour after operation were compared and revealed no statistical significance. The scores of Wong-Baker and FLACC in children at 6 hours and 24 hours postoperative were compared subjectively and objectively and revealed statistic significance. Conclusions Bibliotherapy can ameliorate the anxiety level of both children and their care givers, relieve the perception of postoperative pain in children and improve their comforts. Bibliotherapy thus conduces to the recovery of postoperative children.

Objective To investigate the eye-movement features of smooth pursuit in subjects at clinical high risk for psychosis.Methods sixty subjects at clinical high risk for psychosis and sixty healthy controls were recruited.The smooth pursuit tasks were assessed in both horizontal (0.4 Hz) and Lissajous (0.2 or 0.4 Hz) condition.The Wechsler Memory Scale-third edition and spatial span subtest were used to assess working memory.The difference of the smooth pursuit performance between the two groups and the relationship between smooth pursuit and working memory were analyzed.Results Subjects at clinical high risk for psychosis showed significantly lower Horizontal components for pursuit gain [Lissajous 0.2 Hz task (0.82±0.12) vs.(0.89±0.09),Lissajous 0.4 Hz task (0.78±0.13) vs.(0.84±0.14)],lower vertical components for pursuit gain [Lissajous 0.2 Hz task (0.80±0.14) vs.(0.86±0.12),Lissajous 0.4 Hz task (0.71±0.15)vs.(0.77±0.16)] and higher mean positional error [Lissajous 0.2 Hz task (37.00±19.10) vs.(30.45± 16.18),Lissajous 0.4 Hz task (44.18±19.70) vs.(37.61±16.26)] compared to healthy controls (P＜0.05).There was a significant correlation between pursuit gain and performance on Spatial Span (Horizontal components:r=0.361,P=0.005;vertical components:r=0.327,P=0.01 1) in the Subjects at clinical high risk for psychosis.Conclusions Subjects at clinical high risk for psychosis showed deficits in smooth pursuit,and the deficits were related to the working memory.

Objective To study the association of glucagon like peptide 1 receptor (GLP1R) gene polymorphism with type 2 diabetes in Han population in Shanghai. Methods In the study, 360 type 2 diabetic patients and 313 normal control subjects were enrolled. Diabetic patients were further subdivided into insulin treated non obese patients (BMI28, 192 subjects). A single nucleotide polymorphism (SNP) rs 2268657 was genotyped in all the subjects enrolled in the study using allele specific real time PCR and its association with type 2 diabetes was examined. Results The frequencies of AA,AG, GG genotype incontrol group were0.086,0.447, 0.466 respectively, 0.155, 0.375, 0.470 in non obese diabetic patient group respectively, and 0.109, 0.500, 0.391 in obese diabetic patient group respectively. There was significant difference of the frequency of genotype AA between control group and non obese diabetic patient group (OR=1.939, P

To produce recombinant Maxadilan using gene engineering technology, the gene of recombinant Maxadilan which expressed in protocaryon were designed and synthesized according to the amino acid sequences of Maxadilan. The recombinant plasmid pKYB-MAX was constructed and transformed into host bacteria Escherichia coli strain ER2566. After the MAX-intein-CBD fusion protein was purified by chintin-affinity chromatography, the self-cleavage activity of the intein was induced by beta-mercaptoethanol and the recombinant Maxadilan was released from the chitin-bound intein tag. The molecular weight of peptides was determined by the laser flight mass spectrometry and the results was conformity with the theoretical value. The biological activity analysis showed that recombinant Maxadilan significantly enhanced the concentration of serum glucose.
Animals ; Base Sequence ; Escherichia coli ; genetics ; metabolism ; Insect Proteins ; biosynthesis ; genetics ; Inteins ; genetics ; Isopropyl Thiogalactoside ; pharmacology ; Molecular Sequence Data ; Recombinant Proteins ; analysis ; biosynthesis ; genetics